When devices for amplification of nucleic acids using the polymerase chain reaction (“PCR”) were first developed, it was necessary to allow the amplification reaction to occur for one cycle at a temperature of approximately 37° C.; to then separate the resulting double stranded DNA by heating to approximately 100° C.; and finally to cool the mixture of separated DNA strands and add more polymerase to cause another cycle of amplification to take place. It soon became clear that these devices were cumbersome to use due to the constant need to cool the individual reaction mixtures to about 37° C. and add fresh polymerase enzyme, due to its destruction when the temperature had been raised to about 100° C.
The advent of thermostable nucleic acid polymerases changed the situation dramatically. These enzymes are stable at about 95° C. and therefore it was not necessary to replace these enzymes after heating to separate the two strands of DNA. Thus, devices which took advantage of a thermostable nucleic acid polymerase were developed.
Current technology uses 96 and 384 well plates real-time instruments to attain high throughput, coupled with automated robotic systems to load, seal and transfer plates to and from real-time thermal cyclers. Using this technology the throughput is limited by the run time of a 384 well cycler which is approx 90 minutes and so equates to 16 runs in 24 hours at 384 well samples and equals approx 6,000 samples.
These robotic systems are physically large, prone to break down, expensive (about $A240,000 as at 2001) and require constant routine maintenance.
Difficulties encountered with these current devices were in part addressed by the invention described and claimed in U.S. Pat. No. 5,270,183 (U.S. '183) which was developed by one of the present inventors (Corbett, Snr) and two others, Reid and Riggs, the contents of which is incorporated herein by reference. In essence, that invention was directed to the reactants travelling through a continuous tube which was subjected to varying temperatures by coiling the tube around substantially drum shaped bodies held at varying temperatures. In order to prevent cross contamination between samples, the reaction mixture was injected into a stream of carrier fluid which separated separate reaction mixtures and passed through two or three separate heating zones. This arrangement allows sequential processing of a number of samples. The carrier fluid and reaction mixture are immiscible. The result of this is that each sample is cleanly separated from the preceding and following sample by segments of carrier fluid.